Biospecific bimolecular binding reactions - a new ellipsometric method for their detection, quantification and characterization.
A new ellipsometric method for detection, quantification and characterization of bimolecular, specific interactions on solid surfaces, e.g., binding between antigen and antibody and between ligand and receptor, is described. In the method, which we have called diffusion-in-gel (DIG) ellipsometry, one of the binding components is placed in a trough in a gel which has been poured over a solid surface coated with the other binding component. After diffusion, the gel is removed from the surface and ellipsometric measurement of thickness of adsorbed bimolecular layers is performed at different distances from the site of the diffusion trough. Bimolecular binding on the solid surfaces was also studied by wettability determinations with a water condensation technique. Three bimolecular binding systems were studied: bovine serum albumin (BSA)-anti-BSA, ganglioside GM1-cholera toxin, and C-polysaccharide-C-reactive protein. There was no tendency to saturation in the anti-BSA adsorption profile, which was steep with an endpoint thickness of about 16 nm. In contrast, the cholera toxin profile, within a narrow concentration range, rose to a plateau level of about 3 nm thickness of adsorbed cholera toxin. The C-reactive protein profile formed an intermediate pattern. Good agreement was observed between the thickness of the adsorbed ligand layers and wettability as determined by water condensation. Compared with other methods, the DIG ellipsometry technique has several theoretical and practical advantages for the detection and investigation of biospecific bimolecular binding.
D000327:Adsorption; D000818:Animals; D000937:Antigen-Antibody Reactions; D001665:Binding Sites; D002097:C-Reactive Protein / Q000276:immunology; D002417:Cattle; D002772:Cholera Toxin / Q000276:immunology; D005677:G(M1) Ganglioside / Q000276:immunology; D005779:Immunodiffusion; D046911:Macromolecular Substances; D008722:Methods; D011135:Polysaccharides, Bacterial / Q000276:immunology; D011817:Rabbits; D012710:Serum Albumin, Bovine / Q000276:immunology
D046911:Macromolecular Substances; D011135:Polysaccharides, Bacterial; C027158:polysaccharide C-substance (Streptococcus); D012710:Serum Albumin, Bovine; D005677:G(M1) Ganglioside; D002097:C-Reactive Protein; D002772:Cholera Toxin